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. 2021 Nov 23;22(23):12639. doi: 10.3390/ijms222312639

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© 2021 by the authors.

Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/).

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SOT1 directly interacts with CAF1 and CAF2. (a) Yeast two-hybrid assays indicate that SOT1 interacts with CAF1 and CAF2. SOT1 was fused to the BD vector; CAF1 (or CAF2, CRS2, and CFM2) was fused to the AD vector. The ability to grow on SD/-Trp-Leu-His-Ade dropout plates indicates an interaction between the two proteins. (b) Luciferase complementation assay for interactions between SOT1 and CAF1 versus CAF2. The indicated NLuc and CLuc constructs were transiently coexpressed in Nicotiana benthamiana plants through Agrobacterium-mediated transformation, and the luciferase (Luc) activity was measured after 48 h. Error bars indicate SD (n = 4). (c) Coimmunoprecipitation assays showing the interaction of SOT1 with CAF1 or CAF2. 3 × HA-tagged SOT1 and 3 × FLAG-tagged CAF1 (or CAF2) were coexpressed in Arabidopsis protoplasts. Protein complexes were immunoprecipitated (IP) using an α-FLAG antibody. Immunoblot (IB) analysis of the protein presence in the immunoprecipitates using the α-FLAG and α-HA antibodies, respectively. The assay of interaction between SOT1 and Ycf4 was used as a negative control.